This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Forces resulting from acoustic radiation pressure are an effective means to localize particles in an arrangement similar to hydrodynamic focusing without the need for sheath fluids. A recently developed acoustic device that has been proven effective in sheath replacement is the line-driven capillary [G. Goddard, et al., Cytometry 69A, 66-74 (2006)]. It is constructed from a capillary that is driven by a piezoceramic source in line-contact with its outer wall. Vibration of the structure creates a localized pressure node along the central axis where an axial particle trap is formed. The implementation of acoustic particle focusing in flow cytometers will enable new flow cytometry techniques and methods to evolve due to fundamental changes in the way particles are positioned within the sample cell. The advantages of this new type of sample delivery include: i.) longer particle interrogation times without concurrent loss in particle analysis rate, ii.) ability to freely stop and reverse the flow direction without loss of particle alignment for particle reanalysis, iii.) induced particle orientation in the optical scattering plane, and iv.) in-line concentration and size fractionation of dilute samples. This proposal is a tailored implementation of acoustic line-driven capillaries to develop new measurements and in-line sampling methods that are made possible by replacing hydrodynamic sheath flow with acoustically focused particle streams. First, we will develop enhanced detection capabilities that utilize slow-flow, stop-flow, and reverse-flow conditions in an acoustically focused flow chamber. Our applications will include investigation of statistical data enhancements that can be achieved when particle reanalysis is allowed and the flow stream can be reversed to reanalyze particles of high significance. Second, acoustic slow-flow (long transit time) particle delivery will be studied for a system based upon time-resolved luminescence detection using lanthanide chelate probes to provide a means for autofluorescence rejection and increased spectral multiplexing. Third, we will investigate acoustic control of particle orientation in the optical analysis plane to study enhanced scattering signatures of asymmetric particles that are oriented in a repeatable fashion in the optical interrogation region. Finally, we will develop acoustic field-based particle size selection for pre-analysis in-line sample purification and pre-sorting for flow cytometers